thermostable proteases

Kresten kresten at my-deja.com
Sat Jan 29 07:16:19 EST 2000


In article <7opiGDf98QgB-pn2-l4vlFuz544H7 at rnaworld.bio.ukans.edu>,
  PGegen at UKans.nolospamare.edu wrote:
> On Fri, 28 Jan 2000 14:32:02, Kresten <kresten at my-deja.com> wrote:

[snip]

Thanks.

Actually we have quite a lot of PrK lying around since we use it for
other purposes. The only thing I didn't realize was that it kept
activity as such low values of pH. Being a serine protease I thought
that PrK would only be active under alkaline conditions but as far as I
can see from the web-literature PrK is active from pH 4.3 to 12.0
having max activity around pH 8. I'll have to check the literature to
see how much activity is left at eg. 60 degrees and pH 5.

Kresten



>
> You're in luck! Proteinase K, available from many sources, performs
> optimally
> at 65 deg. C. A typical protocol for *complete* proteolysis would be
> 0.1 mg
> proteinase/mL, 0.1-1.0% SDS. (The enzyme also tolerates urea up to 3-4
> M, and
> EDTA to 0.1 M or more.) There isn't much self-digestion at room
> temperature,
> but you should keep the enzyme frozen when not in use. It's a serine
> esterase
>
>
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Kresten Lindorff Larsen, Dept. Yeast Genetics


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